In the test, the technology corrects the genetic defect of a neuromuscular disease without a cure

Since the end of the human genome project at the turn of the 20th and 21st centuries, one of the greatest goals of scientists and doctors has been to use the knowledge gained from the three billion letter pairs in our DNA to treat diseases.

One of them is Duchenne muscular dystrophy (DMD), a serious neuromuscular disorder that usually kills patients in their third decade of life.

Scientists at the University of Texas’ Southwestern Medical Center (UTSW) succeeded in developing gene therapy tools in the laboratory that can permanently correct the most common type of genetic defect of the disease in cells and mice. The research was published on Friday (30) in the journal Science Advances.

The disease affects 1 in 3,500 boys born. The girls are spared because even if they receive a defective gene, they almost always have a good copy on a different X chromosome – one inherited from the father and the other from the mother. Boys, on the other hand, only have one X chromosome inherited from their mother. in them, the opposite sex chromosome is the Y inherited from the father.

This altered gene makes muscle cells extremely fragile. During muscle contraction, they cannot maintain structure, which leads to a kind of self-destruction. In addition to functional disorders, there is inflammation, which also leads to muscle disorders.

Not only are the muscles associated with the skeleton (such as biceps, calves, pecs, glutes, quadriceps, etc.) affected, but the heart as well. The body cannot repair the muscles because they are destroyed. Both the heart and respiratory functions are impaired.

For the study, the scientists made a mouse that showed the same type of change as 8% of those with DMD. This type of change, as in most cases of the disease, causes a DNA read error, which results in the dystrophin protein coming out incomplete and inoperable.

With this in mind, the researchers developed a plan to remedy this deficiency, but it was not a trivial task. What they managed to do was rearrange the exons, which are the DNA sequences that make up a piece of the protein. There are 79 exons in the dystrophin gene, which means we can think that the protein has 79 pieces in a predefined sequence.

The defect in question is the deletion of exon 51. The challenge was to change the way the sequence is read and definitely translated into a protein. For this it was necessary to save everything that came from exon 52.

By changing the letter (or base) at the end of exon 50, it was possible to have the reading jump from exon 49 to exon 52, creating a slightly shorter version of dystrophin, but which was able to perform most of its cellular functions .

A basic editing technique was used for this (changing “lower case letters”). The studies were carried out on mice and on heart muscle cells (cardiomyocytes) cultivated in the laboratory.

Another tactic was to add two bases to exon 52, which prevented the protein from shortening. This was done in heart cells based on a probe (which is responsible for finding the right place to change the DNA molecule) and molecular scissors that were able to make the desired changes.

In both experiments with cultured cells, not only did detection of the protein begin, but there was also a reduction in arrhythmia, one of the signs presented by the defective cells.
“In principle, completely restoring the missing exon would be the best strategy, but there are currently no gene-editing technologies available to do this in muscle fibers and cardiomyocytes,” said study lead author Francesco Chemello and senior author Eric Olson by email.

According to the neurologist Sarah Camargos, professor at the UFMG, the new work brings optimism into the fight against Duchenne’s dystrophy. “With this finer output and this exchange of bases, a world of mutations can be tackled,” he says. “It’s an example of personalized medicine that can treat multiple deletions in the same gene.”

Jonas Saute, coordinator of the outpatient clinic for genetics of neuromuscular diseases at the Hospital de Clínicas de Porto Alegre and professor at UFRGS, explains that other attempts at gene therapy for DMD have already been made, for example with the introduction of microdistrophin and mini-distrophin – much shorter versions of dystrophin, but which theoretically still retain a certain function. However, the results are still not as consistent.

Currently, according to Saute, treatment is available to patients with corticosteroids (anti-inflammatory agents) and some molecules that attempt to “force exon skipping” with the disadvantage that it infuses all week Need to become.

“We are working to optimize these strategies and achieve the best correction efficiencies for the most common mutations in patients with DMD. Unfortunately, there are some less common mutations in certain essential regions at the beginning and at the end of the DMD gene that cannot be manipulated this way. One of our future goals is to find a solution to correct these mutations as well, ”the authors of the study say of the report.

One obstacle to human studies, according to researchers, is the possibility of genetic changes in the wrong places (and potentially catastrophic effects, such as cancer tendencies). Chemello and colleagues did not find any nonspecific changes, but this is an analysis that must be performed for any type of deletion in any attempt at treatment.

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